Holzgrabe group: research topics

Medicinal Chemistry

"Medicinal chemistry is a chemistry-based discipline, also involving aspects of biological, medical and pharmaceutical sciences. It is concerned with the invention, discovery, design, identification and preparation of biologically active compounds, the study of their metabolism, the interpretation of their mode of action at the molecular level and the construction of structure-activity relationships." (http://www.chem.qmul.ac.uk/iupac/medchem/ix.html#m1)

We focus on a variety of topics in the field of medicinal chemistry and pharmaceutical analysis, including:

Small molecules against infectious diseases

These projects are linked to the Collaborative research center (SFB 630): Recognition, Preparation and Functional Analysis of Agents against Infectious Diseases (see: http://www.sfb630.de/).

Smart and small molecules are aimed to be developed for treatment of tuberculosis, legionellosis, African sleeping, sickness and leishmaniosis. Both ligand- and structure-based drug design will guide the way to innovative active substances.

Quinolone-carboxamides against Trypanosoma brucei brucei

The mitochondrium change from an elongated tubular shape to sheet-like patches. Other organelles are not affected ( Taken from J. Med. Chem. 55, 2538-2548 (2012))

By serendipity, the high activity of the quinolone-carboxamides against Trypanosoma brucei was detected and optimized. The best compound is shown here. Mode of action: change in the morphology of the mitochondrium; most likely the defect in kinetoplast segregation. This mechanism is not identical with ciprofloxacin and thus, the target is not necessarily the topoisomerase.

Lipophilicity logP 2.4 to 4.0

Mol. Weight 400 to 550

Less than 10 H-bond acceptors

Less than 5 H-bond donors

Low water solubility

IC50 (T.b.brucei, 72h) = 47 nM

IC50 (BSF T.b.b., 48h) = 23 nM

IC50 (T.b. rhodesiense, 72h) = 9 nM

IC50 (J774.1 macrophages) = 57 µM

Cytotoxicity:The higher trypanocidal activity, the more pronounced the gap to cytotoxicity:SI > 1000 can be easily achieved

Tertiary bisammonium bisnaphthalimides are known to be cancerostatic. However, the quaternary analogues do not exhibit this cytotoxicity, but are active against tropical protozoa and staphylococci. Of note, the structure-activity relationships of the naphthalamides are different in the different protozoa and Staphylococcus aureus.

Aims: Optimization of the compounds against the various microorganisms. Bisquaternary bistacrine derivatives show a similar activity. Optimization of this series with regard to antiinfective activity and toxicity.

KasA inhibitors against Mycobacterium tuberculosis

The structures of the human and bacterial enzymes building fatty acids are already elucidated. This holds true inter alia for the elongation-condensation enzyme FabB of E. coli. The corresponding enzyme KasA was isolated from mycobacteria and structurally characterized in presence of thiolactomycin inhibitor. Virtual screening gave suggestions for inhibitors which are synthesized and biologically evaluated by means of a fluorescence assay. Some compounds exhibiting good KasA inhibitory activity were identified which are in part active against mycobacterium tuberculosis.

Aims: Development of an enzyme assay which specifically measures the inhibition of KasA; crystal structure of KasA complexed with a newly synthesized inhibitor.

Development of anticancer drugs active against multiple myelom

This project is linked to the clinical research unit (CRU 216): "Characterization of the Oncogenic Signaling Network in Multiple Myeloma: Development of Targeted Therapies" (for more information click here ).

Inhibitors of the HSP70 chaperone (structure-based desgin)

Heat shock proteins 70 (Hsp70) are highly conserved molecular chaperones, ubiquitously present in eukaryotic cells. Due to their numerous cellular functions, including correct folding and refolding of proteins, prevention and dissolution of protein complexes, degradation of unstable and misfolded proteins, and the control of regulatory proteins, these enzymes play an important role in protein homeostasis. The family of 70kD chaperones comprises two kinds of proteins, stress-inducible and constitutively expressed isoforms, also known as heat shock cognates (Hsc70). In recent years, increasing evidence has suggested Hsp70 as a potential anti-cancer target. In this project we target the interface between the substrate-binding protein and the nucleotide binding site. By means of virtual screening a hit was found and the correspondingly developed compound library led to a highly active inhibitors.

Aims: Further optimization of the inhibitors, elucidation of the molecular mode of action; development of inhibitors of the HSF-1 protein.

Dualsteric ligands of muscarinic receptors

Since a long time we are interested in allosteric modulators of muscarinic receptor. In collaboration with K. Mohr (Bonn) we have developed positive allosteric modulators and have taken the concept to bitopic dualsteric compounds which simultaneously bind to the orthosteric and allosteric site of the muscarinic receptor. Beside receptor subtype selectivity a signalling selectivity was observed.

Pharmaceutical quality analysis

The department has a long standing tradition in the development of monographs for the German Pharmacopoeia (Deutsches Arzneibuch) and the European Pharmacopoeia. This is especially true when it comes to the identification, separation, and quantification of the impurity profile of drugs. Employing HPLC-UV; HPLC-CAD, HPLC-ELSD, as well as capillary electrophoresis and sometimes NMR spectroscopy the quality of the drugs can be ensured.Capillary electrophoresis techniques (CZE, MEKC, MEEKC) modified with various cyclodextrin derivatives are often used for chiral analysis. NMR spectroscopy and molecular simulations were employed for the elucidation of the mode of chiral recognition.

Quantitative NMR spectroscopy

qNMR spectroscopy can be used as an orthogonal method for the quantification of impurity in a drug. Parameter setting for measurement and processing is critical for sensitivity and precision of the method. Beside quality assurance of drugs, qNMR is utilized in unraveling counterfeits.Different NMR techniques, such as DOSY, ROESY, etc., are used for elucidation of counterfeits from drug formulations, for characterization of e.g. ionic liquids and others.

Counterfeit medicines

In the beginning of the century we were involved in the gentamicin case. Some 80 people died in the US according to impure gentamicin. Using MEKC and qNMR spectroscopy we could characterize the impurity profile of the dangerous batches. More recently we were involved in the heparin case where heparin was poisoned with oversulfated chondroitin sulfate. We have observed the German market by using qNMR and CZE. In addition biological assays were employed in collaboration with S. Alban (Kiel) and B. Wolff (Ratiopharm, Ulm).